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Combining 3D Raman Imaging And Surface Structural Imaging

Harald Fischer, Sales and Marketing manager at WITec, talks AZoM about the benefits of 3D Raman imaging and how this can be combined with surface structural imaging techniques.

Could you please provide a brief introduction to 3D Raman imaging and how this is commonly utilised?

For 3D Raman Imaging a Raman spectrometer is attached to a confocal microscope, so that the resolution of the optical microscope is combined with the analytical power of Raman spectroscopy. In Raman spectroscopy, a vibrational quantum state is excited or annihilated within a molecule, leading to an energy shift between the incident light and the scattered light. This energy shift is unique to each molecule and allows the chemical identification of compounds within a sample.

To generate images with the confocal Raman microscope, the sample is scanned point-by-point and line-by-line, and at every image pixel a complete Raman spectrum is taken in a continuous readout process. This procedure is also called hyperspectral imaging.

These multi-spectrum files are then analyzed to display the distribution of chemical sample properties. By taking a stack of images with different focal positions, the geometry of samples can be reconstructed in 3D with a lateral resolution down to approx. 200 nm in xy and 500 nm in the z direction.

Combined Raman-AFM imaging system alpha300 AR for high resolution structural and chemical imaging.

Combined Raman-AFM imaging system alpha300 AR for high resolution structural and chemical imaging. Image credit: WITec

How quickly can a Raman spectrum be collected and what can be determined from the resulting image?

Typical integration times in a confocal Raman microscope are between 700µs and 100ms per spectrum and pixel, so that a complete Raman image of 10,000 spectra or more takes between a few seconds or minutes. We established the term ultrafast Raman imaging for this kind of measurements.

The resulting images not only reveal optical information but also information regarding the 3D distribution of the chemical compounds, crystallinity and material stress.

When would Tip-Enhanced Raman Spectroscopy (TERS) be applied and what are the benefits of this?

According to Abbe’s theory of diffraction and Rayleigh's definition of the diffraction limit, a spatial resolution of approx. 200 nm can be achieved with confocal Raman imaging. As an alternative approach to achieve lateral resolution far below the diffraction limit Tip- Enhanced Raman Spectroscopy can be applied.

In a typical TERS experiment an Au or Ag-coated AFM tip is used as a nanostructure to produce Raman signal enhancement on a sample surface once the excitation laser is focused on the apex of the tip with the tip brought into close proximity with the surface. The tip radius, which defines the lateral resolution of an AFM measurement, is typically in the range of 10 – 20 nm.

In a TERS experiment the lateral resolution depends on the size of the hot-spot therefore one can expect resolution in the range of 20-50 nm for Raman spectroscopy and imaging measurements which is a mayor advancement. Microscopes with integrated Raman-AFM capabilities such as the WITec alpha300 series are therefore ideally suited for TERS experiments.

Additionally it is possible to perform near-field Raman imaging using an aperture SNOM system delivering Raman images with an optical resolution beyond the diffraction limit paired with the ease of use and reliability of the established WITec cantilever SNOM-probe technique.

What are the major analytical benefits of combining Raman imaging with techniques such as AFM or profilometry?

Combined analytical microscopes allow a direct linking between high resolution surface structural imaging and chemical identification of various species on a surface.

For example a topographic defect on a samples surface can be immediately be associated with chemical properties contributing to a more detailed and comprehensive understanding of your micro- and nanostructures.

How can this be utilised in materials science research, such as thin film analysis?

Coatings and films play an important role in many fields of application which makes an efficient technique for the investigation of these materials essential. The combination of Raman imaging with AFM can be beneficial for the nondestructive characterization of such coatings in a sense that Raman delivers information on the distribution of chemically different materials and AFM reveals the topographic structures of the films with a resolution down to a few nanometers.

Detailed information on the 3D structure of multi-layered polymer films or coatings can be attained using the depth profiling capabilities of the confocal Raman microscope. Both techniques are nondestructive and require minimal sample preparation if any.

Raman images showing the distribution of the two polymers.

Raman images showing the distribution of the two polymers. Image credit: WITec

In materials research both techniques are very well established for the analysis of nano-carbon materials such as carbon nanotubes or graphene. Graphene consists of carbon atoms which form angstrom-thick two dimensional sheets.

These sheets occur as multilayers in graphene flakes. While AFM can provide information about the physical dimensions of nano-materials, Raman imaging gives insights into the molecular composition of a material.

Using the combination of confocal Raman microscopy with AFM, the high spatial and topographical resolution obtained with an AFM can be directly linked to the molecular information provided by confocal Raman spectroscopy.

How can this be applied in other areas, such as the mining and pharmaceutical industry?

The geosciences and mining community requires distinct information on the appearance of certain minerals within rock sections or tiny dust particles.

A Raman image with corresponding spectra allow for the general assignment of mineral phases and their gross distribution over a scanned area.

In addition to the mineralogical context information, organic components can be identified, spectrally characterized and located if trapped between precipitates.

3D Raman image of a fluid inclusion (blue) in garnet (red).

3D Raman image of a fluid inclusion (blue) in garnet (red). Image credit: WITec

In the pharmaceutical industry microscopes are used primarily for the characterization of drug delivery systems or medical devices.

The surface structures of such devices can be recorded and matched with the drug distribution, thickness parameters or homogeneity of the drug delivery materials. Even polymorphic variations can be easily distinguished.

Systematic and routine research tasks with repetitive experiments or a large number of measurement points, as well as high-level quality control can benefit from automated instrument functionalities.

AFM (phase) image of the surface of a drug delivery coating.

AFM (phase) image of the surface of a drug delivery coating. Image credit: WITec

What are the benefits of simultaneous cantilever and sample observations achievable using WITec's alpha300 A microscope?

With its integrated research grade optical microscope, the alpha300 A provides high-resolution and simultaneous sample and cantilever survey from above.

This makes sample positioning very easy and the alignment is straightforward when, for example positioning the AFM tip accurately on very small sample structures.

The result is an extremely user-friendly instrument for materials research, Nanotechnology and Life Sciences supporting all standard AFM modes.

Focussing on the alpha300 R, why is it useful to have resolution down to the optical diffraction limit?

As the trend towards miniaturization continues it is important to have imaging tools that deliver the most advanced spatial resolution.

The highest lateral resolution that can be achieved with an optical microscope is described by diffraction theory and depends on the excitation wavelength and the numerical aperture of the objective used for image generation.

As a rule of thumb one can consider half of the excitation wavelength as an approximate value for the achievable optical resolution.

It should certainly not be limited by any components or the design of the instrument itself but rather only by the laws of physics.

The optical design of the WITec microscopes always allows for diffraction-limited and ultrafast 3D Raman imaging to be performed with one and the same instrument configuration routinely and simultaneously.

The main parameters we have defined here are “Speed”, “Sensitivity” and “Resolution”. From our point of view, it is important to fulfill all three parameters and not just one or two as scientists typically want to rely on cutting-edge tools in order to set the benchmark in their field of application.

Could you explain the principles behind WITec´s TrueSurface Microscopy?

WITec’s exclusive True Surface Microscopy mode makes it possible to perform confocal imaging measurements parallel with and guided by large area topographic scans (> 1x1mm²). To achieve this unique capability, the WITec microscope series can be equipped with a highly precise sensor for optical profilometry.

The large area topographic coordinates from the profilometer measurement are used to perfectly trace the samples surface in either confocal or confocal Raman imaging mode. Sample preparation is reduced to a minimum without having to compromise the confocality of the system.

The TrueSurface Microscopy mode is beneficial for many applications, including the characterization of micromechanical, medical, or semiconductor devices, the mapping of functionalized surfaces, or the imaging of bio-medical or pharmaceutical material surface properties.

How do you see the combination of 3D Raman imaging with surface structural imaging techniques progressing in the future?

In general there will be further improvements in sensitivity along with new developments in detector technology. Another element of successful instruments will be an improved hardware and software user interface and user-friendly data evaluation for automated post-processing of the Raman spectral data.

With regard to combined instrument configurations there is an ongoing trend toward correlative microscopy and the users can definitely expect more in that direction from WITec.

About Harald Fischer

Harald Fischer

With a scientific background in Biology and Chemistry, Fischer started his career in the IT industry holding positions as a marketing and product marketing manager. In 2002 he joined WITec, a German manufacturer of nano-analytical microscope systems, as a Sales and Marketing manager.

He soon became responsible for the company’s worldwide marketing-communication activities and is now working as Marketing Director leading the WITec marketing team at the headquarters in Ulm, Germany.







Disclaimer: The views expressed here are those of the interviewee and do not necessarily represent the views of Limited (T/A) AZoNetwork, the owner and operator of this website. This disclaimer forms part of the Terms and Conditions of use of this website.

G.P. Thomas

Written by

G.P. Thomas

Gary graduated from the University of Manchester with a first-class honours degree in Geochemistry and a Masters in Earth Sciences. After working in the Australian mining industry, Gary decided to hang up his geology boots and turn his hand to writing. When he isn't developing topical and informative content, Gary can usually be found playing his beloved guitar, or watching Aston Villa FC snatch defeat from the jaws of victory.


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